Down syndrome (DS) children have a high risk of developing acute myeloid leukemia (AML). AML in DS has unique features: a) it typically presents with a megakaryocytic phenotype (AMkL);b) event-free survival rates for DS-AMkL are much higher than non-DS children with AMkL (~75-100% versus <25%);c) AMkL may be preceded by a transient myeloproliferative disorder (TMD) which occurs in approximately 10% of DS newborns and evolves into AMkL in ~20%, while resolving spontaneously in the others. The molecular and cellular events underlying the unique pathobiology of AML in DS remain to be elucidated. Small retrospective studies have identified somatic mutations of the X-linked transcription factor gene, GATA1, exclusively in DS TMD and AMkL cases. Conceivable, GATA1 mutations could contribute to leukemogenesis and also enhance the sensitivity of leukemia cells to chemotherapy. This proposal takes advantage of correlative biological studies planned in the Children's Oncology Group clinical trials AAML0431 "Treatment of DS Children with AML and MDS Under the Age of 4 Years" and AAML0532 "The Treatment of DS Children with TMD". Studies in Specific Aim 1 will determine and compare the types of frequency of GATA1 mutations in DS TMD and AML cases and their relationship to the AMkL phenotype. The identification of secondary genetic events linked to the development of AMkL including JAK3 mutations and altered microRNA expression will also be pursued in this Aim. In Specific Aim 2, microarray studies will identify genes that predict TMD outcome, and to describe the molecular events associated with the transition from TMD to AMkL. In Specific Aim 3, pharmacologic and molecular assays to study the response of DS AML cases to therapy including the use of minimal residual disease (MRD) detection to monitor treatment response during induction therapy, which may be used to identify the appropriate cytarabine dose in clinical trials. Within this aim, a novel molecular assay for MRD detection targeting GATA1 mutations will be developed and the results validated by comparisons with those obtained with the established flow cytometric MRD assay. Success in the studies proposed under these aims should lead to improvements in the clinical management of children with DS and AML, and also provide unique information about the fundamental mechanism that regulate leukemogenesis and drug sensitivity in AML.